Abstract
The ATP-sensitive K(+) (K(ATP)) channel is a complex of a pore-forming inwardly rectifying K(+) channel (Kir6.2) and a sulphonylurea receptor (SUR). The aim of the present study was to gain further insight into the mechanism of block of K(ATP) channels by terfenadine. Channel activity was recorded both from native K(ATP) channels from the clonal insulinoma cell line RINm5F and from a C-terminal truncated form of Kir6.2 (Kir6.2Delta26), which - in contrast to Kir6.2 - expresses independently of SUR. Kir6.2Delta26 channels were expressed in COS-7 cells, and enhanced green fluorescent protein (EGFP) cDNA was used as a reporter gene. EGFP fluorescence was visualized by a laser scanning confocal microscope. Terfenadine applied to the cytoplasmic side of inside-out membrane patches concentration-dependently blocked both native K(ATP) channel and Kir6.2Delta26 channel activity, and the following values were calculated for IC(50) (the terfenadine concentration causing half-maximal inhibition) and n (the Hill coefficient): 1.2 microM and 0.7 for native K(ATP) channels, 3.0 microM and 1.0 for Kir6. 2Delta26 channels. Terfenadine had no effect on slope conductance of either native K(ATP) channels or Kir6.2Delta26 channels. Intraburst kinetics of Kir6.2Delta26 channels were not markedly affected by terfenadine and, therefore, terfenadine acts as a slow channel blocker on Kir6.2Delta26 channels. Terfenadine-induced block of Kir6. 2Delta26 channels demonstrated no marked voltage dependence, and lowering the intracellular pH to 6.5 potentiated the inhibition of Kir6.2Delta26 channels by terfenadine. These observations indicate that terfenadine blocks pancreatic B-cell K(ATP) channels via binding to the cytoplasmic side of the pore-forming subunit. The presence of the pancreatic SUR1 has a small, but significant enhancing effect on the potency of terfenadine.